Euryodendron excelsum is a critically endangered Ternstroemiaceae species endemic to southern China, with only 14 individuals surviving in Ba Jia Zhen of Yangchun, Guangdong Province. Intersimple sequence repeat (ISSR) markers were used to assess genetic variation and relationships among these individuals. Population genetic parameters were estimated by a Bayesian approach as well as conventional methods. Of the 225 loci generated by 21 primers, 147 (65.33%) were polymorphic. Compared with other species of Theaceae and related families, a high level of genetic variation was identified in E. excelsum (Nei's gene diversity, 0.2458; Shannon's index, 0.3626). An unweighted pair group method with arithmetic mean (UPGMA) dendrogram showed that the 14 individuals were mainly clustered into three groups, a conclusion further supported by principal coordinate analysis. Based on these results, a management and conservation strategy for E. excelsum was proposed.